The transdermal use of cyanobacterial algae, and/or extracts thereof, via low or high intensity transporation devices for beauty and health enhancing purposes

ABSTRACT

The present invention refers to the transporation, or force-transdermal vehiculation, of cyanobacterial algae or extracts thereof, through electroporation, sonoporation (sound), photoporation (light), radioporation (radio frequency), and through any other transporation tool for enhancing the transdermal penetration of cyanobacterial molecules via the transmission of some force, be it electric, or generated by sound, light and/or radio frequencies. The invention cover the use of whole cyanobacterial algae or extracts thereof, with different levels of transdermal penetration, be it the low intensity transporation, which can be used for deep aesthetic and skin regenerating purposes, as well as dermatological problems; or the high intensity transporation, able to penetrate deep though the dermis, and beyond the hypodermis into organs and tissues, and that can be uses for muscular, osteo-articular and metabolic problems. Following an exhaustive explanation of the invention, we present very significant results in different beauty and health areas.

BACKGROUND OF INVENTION

The idea of inserting a substance in a specific part of the human body for therapeutic purposes starts with the invention of the syringe with a needle by the French surgeon Pravaz in 1840. Afterwards, there have been many researches and attempts to find ways that could be less traumatic and more efficient. One such system, for instance, is mesotherapy, which still utilizes the syringe idea, but uses a certain number of micro-needles. The first non-syringe related transdermal technology has been ionophoresis, which uses an electrical continuous current to administer transdermally ions of different pharmaceutical substances. This original method has then been improved upon by such method as iontophoresis, a form of electroporation; or even ultrasound technology, which uses variable frequencies ultrasounds to create an energy impulse capable of opening up the outer strata of the skin, helping molecules to go through. Recent studies have shown for instance how iontophoresis has the same ability to promote the passage of high molecular weight substances through the human sclera (Nicoli S. et al, 2009), while the same has been claimed for systems combining different methodologies (Mitragotri S., 2000).

Different methodologies and equipment can be used to push whole molecules, even large, through the skin or the dermis, down to even tissues and organs, and such system have expanded to include also radio frequency (RF); vibration and cavitation by sound; phototherapy by laser or LED light; and even the application of thermal energy. However, the aesthetic or therapeutic result of such enterprises do not depend solely or principally on the technique used, but preeminently on the type of molecule used and on its therapeutic effectiveness.

Cyanobacteria algae are the source of many both nutritional and nutraceutical molecules, endowed with antioxidant anti-inflammatory, anti-proliferative, neuromodulating, neuroprotective, immune-enhancing and so on (as we shall see in the course of the presentation). We have already developed two patents on extracts from Klamath AFA algae, one on its neurologically active molecules (WO2008/000430), and another one on its special phycocyanins and their antioxidant, anti-inflammatory and anti-proliferative use (WO2008/000431). However, none of those patents, nor other patents, have ever considered the transdermal use of the whole cyanobacterial algae or of their phycocyanins, or of other antioxidant, anti-inflammatory and anti-pain molecules, such as phenylethylamine (Mosnaim A D et al. 2014), contained in them.

In our novel experimentation, we have found for instance that the transdermal use of the Klamath microalgae and of its phycocyanins extract, alone or in synergy with others active ingredients (including Spirulina's phycocyanins extract), generates better and faster results than those obtained using other and possibly more well-known nutritional and pharmacological molecules, as will be shown in the next clinical and factual examples.

THE INVENTION

The invention refers to specific ways to transdermally vehiculate cyanobacterial algae or extracts thereof, such as through electroporation, sonoporation (sound), photoporation (light), radioporation (radio frequency) cavitation, and any other transporation tool which can enhance the transdermal penetration of cyanobacterial molecules. Transporation is achieved via the transmission of some force, be it electric, or generated by sound, light and radio frequencies. These systems, which differ from generic transdermal patches, which are based simply in prolonged contact, use some type of force, be it electrical or other, to promote deeper skin penetration by opening the water and ionic channels within and outside the skin's cells. Such systems can also modulate, via the intensity of the force and the time of the application, the depth of the penetration, achieving a partial penetration of the substance vehiculated down to the dermis, to affect for instance elastin and collagen fibers when using low level transporation intensity or shorter times; while reaching down trough the dermis and into the hypodermis, to affect organs and tissues, via higher intensity and longer time transporation. In this way, these transporation devices allow for the penetration of high molecular weight cyanobacterial molecules such as phycocyanins or large cyanobacterial carotenoids, in order to alleviate and possibly resolve both superficial, epidermis-related beauty and dermatological problems, and deep-rooted skin problems, as well as muscular and bone affections, and possibly even metabolic and organ pathologies.

If we consider for instance phycocyanins, the typical cyanobacterial molecules that have known antioxidant (Bath V. B. et al. 2000; Benedetti S. et al. 2004; 2010), anti-inflammatory (Hsiao G et al., 2005; Romay C. et al, 1998) and anti-proliferative (Scoglio S. et al, 2016) properties, when applied cosmetically, or even applied through transdermal patches, they can only operate superficially. What makes transporation different from mere cosmetics cremes is that it allows for the penetration of the epidermis down to the dermis, to affect for instance elastin and collagen fibers, and even through the dermis and into the hypodermis with enough concentration as to effectively affect muscles, bones or organs. Our previous patent (PCT/EP 2007 005623/published as WO2008/000431) also covered the cosmetic use of AFA-phycocyanins (the name we use for the specific phycocyanins of Klamath microalgae). However, through cosmetic cremes or transdermal means such as transdermal patches, phycocyanins could not go very deep through the skin, and could have mere cosmetic effects; while with transporation, at low intensity levels they can reach the dermis, affecting elastin, collagen and the connective tissue supporting the skin; while with high intensity transporation devices they can reach through the dermis and, via the hypodermis, the underlying organs and tissues, such as muscle, bones, and the bloodstream.

The fundamental problem solved by such transdermal means it's bypassing the resistance of the stratum corneum of the our skin: the phospholipidic intercellular belayer of the skin is naturally hydrophobic; and only the long standing research that led to the discoveries by MacKinnon and Agre (Nobel prize in Chemistry in 1993—FIG. 1), showed that cells have specific and selective channels that allow for the passage of both water and ionic salts. Essentially both low and high intensity transporation work on the same principle of opening up the water and ionic channel in the lipid hydrophobic layer, their difference being only in terms of the depth of penetration of the molecules into and through the skin.

As the process consists in the temporary stimulation of an existing physiological structure and function, the process itself is absolutely physiological, and thus generally safe. However, it is a form of artificial modification of the state of the epidermic cells's water and ionic channels, and this is what distinguishes transporation methods, in its different forms (electro, sono, photo, RF, thermal), from mere transdermal cosmetic means, such as transdermal patches, which do not modify the status of water and ionic channels of the skin's cells, and are thus limited to the transdermal vehiculation of only small molecules, characterized by a molecule size and weight small enough so that the skin does not constitute a barrier for them. So, for instance, molecules such as phycocyanins, which are quite large, or even whole cyanobacterial algae, are too big to penetrate into and through the dermis by themselves.

Once applied through trans-poration methods, phycocyanins, as well as large anti-oxidant and anti-inflammatory molecules contained in the microalgae, such as the AFA-phytochrome described in the above mentioned patent (WO2008/000431) with a molecular weight of approximately 400 kD, can directly and positively affect deep aesthetic and dermatological, as well as muscular, osteo-articular, circulatory and even metabolic diseases, thus bypassing the issue of metabolization through the digestive tract, and throughout the circulatory torrent, which indeed significantly reduces the ability of large, high molecular weight, molecules to arrive to the local, peripheral site where they are needed.

The invention, thus, does not refer to any specific technology or machinery, but to the fact that any result obtainable with the different transporation devices available, is radically enhanced, both in terms of efficacy, speed and cost, by using as the main substance to be vehiculated cyanobacterial algae, such as Klamath algae (Aphanizomenon flos aquae) and/or Spirulina, and/or any of the cyanobacterial antioxidant and anti-inflammatory molecules concentrated in any cyanobacterial extracts.

Other substances can be added to the whole cyanobacteria and/or cyanobacterial extracts, such as phenyethylamine, hyaluronic acid, herbs, specific minerals, enzymes, and so on, but always using the whole cyanobacterial microalgae and extracts as the main component of the products.

The invention consists thus in the transdermal use of a powder, solution, gel or liquid, composed of the whole cyanobacterial algae, be it Klamath AFA, Spirulina or any other, by itself or together with any cyanobacterial extracts concentrating the antioxidant and inflammatory substances such as phycocyanins, phenylethylamine (PEA), MAAs, polyphenols, carotenoids, chlorophyll (Kamat J P et al, 2000; Hartigg U. 1998), etc., to be used trans-topically via different transdermal and trans-poration systems, for health and beauty purposes, through devices that are able to open up the water and ionic channels of the skin. Some of these molecules, such as MAAs and PEA, are quite small, but being embedded in the whole cyanobacterial matrix, they follow the destiny of the larger molecules contained in the matrix.

The preparation can be constituted also by a mix of different cyanobacteria or cyanobacterial extracts, and can be put into bottles, sachets, or any other containers.

Even though the invention implies the possibility of using each cyanobacterial algae and each cyanobacterial extract by itself, the preferred preparation is indeed a mix of both the whole cyanobacterial algae and the cyanobacterial extract, such as the one described in the patent WO2008/000431. This is because, while the cyanobacterial extract concentrates some of the most powerful antioxidant and anti-inflammatory substances contained in the microalgae, the whole microalga itself contains further components, such as vitamins, minerals and other nutraceutical molecules, that can add a significant further action to the vehiculated preparation.

For instance, cyanobacterial algae contain some trace minerals that may not be very effective when ingesting the algae orally, given their dispersion through the overall body metabolism, and the possibility that they may not be able to cross the intestinal membrane; but that, when vehiculated transdermally, they may actually reach the site that needs to be helped and healed much more directly. A relevant example is that of boron, a trace mineral that can be very helpful for the fixation of calcium in the bone, and thus for osteoporosis, and that is present in Klamath algae at a concentration of about 11 mcg./gr., a low quantity in terms of recommended daily intake, and that with the algae taken orally would be relatively ineffective, when we consider that the few micrograms present would be dispersed throughout the body at the level of naonograms, if not picograms. However, even just 11 micrograms, when administered via transporation onto the area affected, can be very relevant and effective.

This is even more true, when we consider that Klamath algae contains about 15 milligrams/gr. of calcium, a dosage that taken orally represent only a small fraction of the daily requirement, but if injected directly into the area covering the affected bone, can actually be very helpful to all osteo-articular and bone problems.

The same can be said of : a) essential fatty acids, such as Omega 3 or Omega 6, present in all cyanobacterial algae, but at a level that, when considered in relation to the oral dose, is still relatively low, whereas it become very effective when applied locally through a transdermal device; b) antioxidant vitamins such as the vitamins C, E and some vitamins B, that can be present in small quantity not very effective hen taken orally, but very useful when injected transdermally; c) many trace minerals present in the algae (AFA algae contains more than 70 minerals and trace minerals), which when taken orally may have a too low concentration to be effective, but when injected transdermally directly in the treated area, they actually can be very effective; d) cyanobacterial algae are also rich in carotenoids, chlorophyll, polysaccharides and polyphenols; but given the low oral dosage (3-5 grams), again their concentration is relatively low (except for a few nutrients), but again it becomes very effective when applied locally, thus without systemic dispersion, via transdermal devices.; e) AFA-algae is also rich in phenylethylamine, a molecule endowed with specific neurological, immunological (Babusyte et al. 2014) and analgesic (Mosnaim A D et al., 2014) properties.

Given the nutritional and nutraceutical richness of cyanobacterial algae, such as Klamath algae or Spirulina, they can generate an optimum synergy with any other phytotherapic and nutraceutical substances. In fact, any single molecule is usually in need of some other molecule playing a supporting role, and in this respect cyanobacterial algae cam provide the best synergy possible. This is why, for each different area of intervention, the cyanobacterial algae, constituting the essential base, can be combined with other substances, each one relevant for the specific area of intervention.

Therefore, the invention also covers not only the transporation vehiculation of cyanobacterial algae and of their antioxidant and anti-inflammatory extracts, but also the synergic use of cyanobacteria with any other substance able to intervene favorable on the specific area to be treated. Such use of the whole cyanobacterial algae and/or their extracts in association with other synergistic ingredients, can be applied to the following areas:

-   A) Aesthetic, including problems such as wrinkles, cellulite,     stretch marks, age spots, muscle and skin tone, breast volume, etc. -   B) Dermatological, including eczema, psoriasis, wounds, scars, etc: -   C) Osteo-articular, including arthritis, inflammation of the joint,     joint calcifications, inflammation and/or calcifications of the     tendons, degeneration of cartilage such as meniscus, osteoporosis,     and so on; -   D) Muscular, such as muscle lesions, traumas, inflammation and     muscle degeneration; -   D) Metabolic, such as lymphatic (diabetes, aging) and circulatory     problems (varicose veins, heavy legs, etc), obesity, fat metabolism,     and so on;

We shall see how the application of products based on Klamath algae and Spirulina as the preferred cyanobacterial algae can have very effective results when associated with specific ingredients in each specific area of application.

Areas of Application in Detail

-   AESTHETICS—In this area we tested the transdermal use, with low     voltage electroporation, as well as low frequency sonoporation and     RF, and low intensity light, of a product composed of the whole     Klamath algae, of Klamath algae's and Spirulina's phycocyanins     extracts, combined with different other substances for each specific     purposes. The main conditions tested are the most difficult to treat     in the field, namely wrinkles, cellulite, stretch marks, tone of     skin, tone and volume of breast, scars and keloids, wounds and so     on; all conditions that do not require any penetration through the     skin and into organs and tissues.

While for tonification and filling purposes, as wrinkles, stretch marks and wounds we only added, to the main cyanobacterial base, high molecular weight hyaluronic acid; for breakdown purposes, in conditions such as cellulite and keloids, we also added some herbal extracts as well as proteolytic and lipolytic enzymes.

-   Wrinkles—Wrinkles are mostly the result of aging, and are due to the     breakdown of collagen emerging with age, and more specifically with     the increase in oxidative and inflammatory processes, of     malnutrition that does not provide the body with enough antioxidants     and protective nutrition (vitamins, minerals, EFA), and a lowering     of the body's ability to regenerate. To respond to this aging     phenomenon, different strategies have been used: providing     antioxidant and anti-inflammatory molecules to the skin; supplying     the skin with missing nutrients; stimulate the skin's ability to     regenerate. Cyanobacterial algae allow all these strategies to be     used at one time: they contain a wide range of bioavailable     nutrients, have a broad spectrum of antioxidant molecules; their     phycocyanins have a high anti-inflammatory power (5); finally,     recent studies have shown that phycocyanins potentially stimulates     the activity of fibroblasts, endogenous molecules that act as     collagen and connective tissue precursors, and so are essential for     skin and dermis to regenerate (9). Thanks to all these properties,     we found that the transdermal application, via low voltage     electroporation of a product containing cyanobacterial algae, the     whole Klamath algae, Klamath and Spirulina extracts, plus high     molecular weight hyaluronic acid, produces profound and rapid     results. Here we report a single case example where, in order to     verify more specifically the ability to work on wrinkles, we decided     to use a high-definition camera to photograph a single very obvious     wrinkle before and after treatment in a woman. [FIG. 2] -   Stretch marks—Stretch marks are a form of scars on the skin, caused     by dermatitis. They can be due to rapid skin stretching associated     with rapid recurrence (common in puberty) or weight gain (i.e.,     pregnancy or muscle mass increase). Stretch marks may also be     affected by hormonal changes associated with puberty, pregnancy, and     muscle mass increase. The main physiological cause of stretch marks     is the fact that glucocorticoid hormones affect the epidermis by     preventing the action of fibroblasts forming collagen and elastin     fibers. This creates a lack of support material, as the skin is     pulled and leads to dermatitis. Phycocyanins have been shown to be     potent fibroblast activity stimulants, and we have already shown how     Aphanizomenon flos aquae (AFA) algae contains a very powerful form     of phycocyanins (6, 7), which is thus preferred, even though it can     be combined for synergic purposes with Spirulina's PCs and other     possible ingredients. Phycocyanins can thus be very beneficial in     treating stretch marks, especially if introduced directly into the     dermis by low intensity electric, sonic photo and RF transdermal     vehiculation. We have experimented with various stretch marks by     applying onto the affected areas the whole AFA algae, an     AFA-phycocyanins as well as Spirulina's PC extracts and high     molecular weight hyaluronic acid, through an     electroporation+sonoporation machine, and remarkable results have     been obtained, as shown in the case reported in FIG. 3; as well as     in the case of a woman of 38 years with obvious stretch marks in her     legs, which have almost disappeared after just 3 treatments (FIG.     4). -   Skin tone—The right skin tone depends on the structural integrity of     the dermis and its essential components, such as collagen and     elastic fibers as well as the underlying connective tissue.     Fibroblasts are precursor cells of all such components, and the     ability to properly activate fibroblasts plays a central role in     maintaining the tone and integrity of the skin. Phycocyanins are     powerful stimulators of fibroblast and fibroblast activity     (Madhyastha, H. K. et al., 2008; 2006) and in that sense seem to be     potentially very useful in restoring and maintaining the correct     skin tone. Our experiences through the transporation of     cyanobacterial algae and phycocyanins extract have confirmed these     hypotheses.

We have tested some cases to see if the transdermal vehiculation of a product comprised of the whole AFA plus AFA-phycocyanins and Spirulina's phycocyanins extracts, plus some high molecular weight hyaluronic acid, can improve skin tone in humans, and the results have bee astounding, as shown in the following cases:

a) The case of a 35-year-old woman whose buttocks showed a tendency towards decreasing dermal tone, which was completely re-established after only 3 treatments (FIG. 5);

b) the case of a 47-year-old man with sagging skin in his arms, whose tone was completely re-established after only three treatments (FIG. 6); c) a 52 year old woman, whose belly's skin was deeply sagging, and whose tone was very significantly restored after only 1 treatment (FIG. 7).

-   Breast tone and volume—The logic behind the effectiveness of the     cyanobacterial transporation in relation to the tone of the female     breast is similar to one already discussed above concerning collagen     and elastin. Regarding breast volume, this may depend on other     factors, which at present are still unknown. Certainly the     fibroblasts stimulation generated by the phycocyanins can play a     role. Though we obtained significant results on breast volume with     the usual product containing whole AFA, AFA and Spirulina's     phycocyanins extracts, as well as high molecular weight hyaluronic     acid, it is possible that adding to the product isoflavones from     vegetable sources, given the hormonal component of breast structure,     could further improve our results. As to breast tone, we report here     one of the most significant cases, a 22-year-old woman with slightly     drooping breasts, whose breasts tone increased very clearly after 4     sessions (FIG. 8). As to breast volume, we report the case of a     young woman with a very small breast, which increased in size quite     significantly after 8 treatments (FIG. 9). -   Cellulite—When we move to cellulite, the situation becomes more     complex, as it involves various factors, such as inflammation, fluid     retention, lymphatic and venous stagnation, hypertrophy of fatty     tissue, with each factor playing a different role in each individual     case.

The product we used in this case, while having at its core still cyanobacterial algae (AFA, Spirulina) and relative extracts, added as complementary ingredients proteolytic and lipolytic enzymes and herbs. The basis for the efficacy of this product is once again based on the antioxidant and anti-inflammatory activity of the active molecules present in cyanobacterial algae. In particular, phycocyanins have been shown to be active also as anti-platelet aggregation factors, and platelet aggregation plays a role in both venous and lymphatic stagnation (8). Also, phycocyanins have shown to have a significant fibrinolytic activity, which is essentially to break down the fibrin that holds up the the cellulite debris together (9). Finally, both proteolytic and lipolytic enzymes are also fibrinolytic, and thus again capable of dissolving the fibrin that holds cellulite tissue together. We have regularly tested the resolution of cellulite cases with the transdermal application of a cyanobacterial based product. Here we present the case of 54-year-old woman with a type 3 cellulite, who received a significant improvement after only three treatments during the same week (FIG. 10). More importantly, we have done a study at a medical center in Italy, in which subjects underwent only 3 transporation treatments, and whose symptoms associated with cellulite were evaluated:

-   -   a) circumference of abdomen and hips, which is known to increase         with the accumulation of cellulite on the tissues (FIG. 11);     -   b) the volume of the legs affected by cellulite, which again is         shown to increase with cellulite and fat deposits (FIG. 12);     -   c) the sense of heaviness and pain associated with the cellulite         (FIG. 13);     -   d) the increase of the diuresis (“aumento diuresi”) associated         with the break down and elimination od cellulite deposits (FIG.         14).

-   Localized adiposity (fat belly and fat thighs)—Localized adiposity     is associated with bad fat metabolism, and fat accumulation in local     tissue, especially around the waist. In addition to the positive     effects that can be created by diet-based systemic intervention,     transporation in the area with excessive adiposity with a     cyanobacterial product, plus certain specific ingredients such as     lipolytic enzymes, can be very effective. Here we have used a higher     intensity transporation, that is higher voltage electroporation. In     fact, fat tissue rests underneath the dermis, and so a deeper     penetration is required to help dissolve and mobilize accumulated     fat. The cyanobacterial algae and extracts seems to be able to     intervene in the synthesis of fat, reducing the production of     arachidonic acid, and hence the inflammation and oxidation of fats     behind the accumulation of localized fat (Kushak R. I. et al.,     2000). Phycocyanins are also accelerators of metabolism, due to     their ability to counter platelet aggregation (Hsiao G. et al,     2005), thus improving blood circulation and ability to remove fat     deposits and debris in various tissues. We tested the transporation     of this formula on various cases of abdominal and thighs fat     accumulation, with rapid and profound results and with obvious     reduction of the belly and legs circumference, as shown in the     following cases:     -   a) a 38-year-old woman who wanted to reduce the circumference         and volume of her thighs, which was visibly achieved, with minus         2.5 centimeters circumference reduction, in only 2 sessions         (FIG. 15);     -   b) a 52-year-old woman, whose belly and waist circumference was         significantly reduced after only 6 treatments (FIG. 16).

-   DERMATOLOGY—In this area we have experienced excellent results     through transdermal application of a product containing     cyanobacterial algae and their anti-inflammatory extracts, in     conditions such as eczema, psoriasis, various forms of dermatitis,     acne, as well as healing wounds and scars.

-   Psoriasis, eczema and acne—The cyanobacterial algae, and especially     the AFA and Spirulina's algae phycocyanins extract, play an     important role here, since all these conditions involve some forms     of skin inflammation. Indeed, in a previous study our research group     showed that even just oral intake of a product containing a     significant amount of AFA-Phycocyanins could produce good results on     various types of psoriasis (Cavalchini & Scoglio, 2009). Low     intensity cyanobacterial transportation has shown to significantly     strengthen this activity. We recorded: a) two cases of localized     psoriasis: in one case, a psoriatic eruption in an area around the     mouth, completely resolved in two sessions; while long-term     psoriasis in the scalp was solved with three sessions; b) two cases     of eczema: in one case, an eczema that afflicted a 16-year-old male     for more than three years was completely removed after only 2     sessions; in another case, a woman with an old-time psoriasis in her     hands had a significant improvement already after a session, and a     complete disappearance of localized psoriasis after 3 more sessions.     Finally, we have also tested two cases of acne: in the first case, a     serious case of facial acne has been considerably improved with 4     sessions; in the second case, a moderate type of acne has been     completely eliminated within 2 sessions. All cases were tested by     transdermal vehiculation of a product containing: Klamath algae and     Spirulina's phycocyanin extract; proteases and lipases.

-   Scars and wounds—Once again, the strong ability of phycocyanins to     stimulate fibroblasts is very important, especially in the healing     of wounds, to the point that they have actually been proposed as     possible remedies for ulcers and other types of wounds (Madhyastha     H K. et al. 2006). We tested a case of diabetic foot ulcer, and with     only three sessions we were actually able to fully heal the chronic     wound (FIG. 17). As for the scars, we have had enough positive cases     with a product that added, to the basic cyanobacterial content (AFA     algae and phycocyanins extract+Spirulina's phycocyanin extract),     also proteolytic and lypolitic enzymes and some special herbal     extracts, such as curcumin and caffeine. Here we present two of     those cases:     -   a) a 30-year-old man with a long, large scar at the right side         of his abdomen had a significant reduction in both visibility         and scar thickness only after four sessions, during which he         also had a parallel reduction in the size and volume of his         belly (FIG. 18);     -   b) a keloid, resulting from a long standing scar, which         significantly and visibly reduced after 5 sessions with the         transdermal application of the above indicated product (FIG.         19).

-   MUSCULAR TRAUMAS, LESIONS AND INFLAMMATIONS—In this area, again,     phycocyanins can play a very important role, both for their     anti-inflammatory and anti-edema power, as well as for their trophic     and regenerative properties, via fibroblasts stimulation. However,     when moving to muscles, we need to use high intensity transporation,     such as higher voltage electroporation or high frequency     sonoporation.

-   Muscle lesions/traumas—With reference to muscle lesions and traumas,     the ability of phycocyanins to inhibit COX-2 and lipoxygenase     enzymes, as well as their ability to modulate factors such as     TNF-alpha, is very important with reference to the need for a rapid     reduction of inflammation and associated pain. Muscle trauma can be     divided into direct (bruises) and indirect injuries (stretches and     tears). The latter are obviously more serious and difficult to cure,     especially if they are 2nd degree injuries. Typical of athletes,     these indirect muscle injuries can hold an athlete of any sporting     activity (or any other human being unable to walk regularly) for     weeks. We have tested four cases of indirect muscle injury:     -   a) the first case concerns a 19-year-old soccer player with an         indirect type 1 injury sustained during the a Sunday game. He         was treated Tuesday and Wednesday and the pain had already gone         away; was then treated again on the following Friday and         Saturday, and the injury appeared completely healed. He did not         play the next game just as a precaution;     -   b) the second case is that of a 45-year-old lady with a chronic         calf inflammation that did not allow her to move regularly. Four         sessions had been scheduled; but after the second session the         calf was already fine. After checking her condition three months         after the second treatment, the lady continued to be well and         had no problems;     -   c) the following two cases refer to two soccer players with a         2nd degree injury; in either case the injury was pronounced. The         two players were treated mid-week for 3 times (Thursday, Friday         and Saturday); by Friday they felt healed, and again they did         not play just our of precaution.

-   OSTEO-ARTICULAR AREA—In the osteo-articular area we are concerned     with problems ranging from arthritis, to inflammation of tendons and     joints, from joints and tendon calcifications to cartilage     degenerations (menus, etc.) to osteoporosis. Once again, the     anti-inflammatory properties of phycocyanins are paramount, and in     cyanobacteria they work in synergy with its other anti-inflammatory     molecules, such as chlorophyll and polyphenols. But the ability of     phycocyanins to stimulate the release and the activity of     fibroblasts, and possibly of other blasts (stem cells), seem to play     an essential role in promoting regeneration of cartilage, bones and     muscles. Here too we need to use high intensity transporation, and     found that such transporation significantly improves the action of     the cyanobacterial extracts, achieving much faster and deeper     results.

-   Inflammation/pain of joints and tendons—We have experienced three     serious cases of inflammation of the joints and tendons:     -   1) a case involving a professional karateka athlete whose knee         was badly compromised and for this reason had to stop training;         the knee was treated twice with the transporation of a product         including cyanobacteria algae and extracts, as well as         proteolytic enzymes and high molecular weight hyaluronic acid,         which adds an acton of its own in the regeneration of         cartilages. After only 2 treatments, the athlete was free of         pain and inflammation, and after 2 more treatments he returned         to training and competing regularly;     -   2) a similar case involved another karateka, which had a wound         to the right tendon and which for this reason had stopped         exercising and competing for a few months. He was subjected to a         treatment with the same product as above. After only three         treatments, the athlete was free of pain and inflammation, and         went back to training and competing regularly;     -   3) an elderly man with a large cyst behind his knee, with pain,         inflammation and inability to completely bend his knee. The man         was treated transdermally with the same product above and         immediately after the first treatment, there was a marked         reduction in the volume of the cyst, which passed from the size         of a big lemon to that os a cherry kernel, plus the complete         disappearance of the pain, and with the recovered ability to         extend almost completely the knee.

-   Joints and tendons calcifications—Calcium deposits tend to form     around the joints and tendons, for reasons that are not too well     understood. At first they are mild, but over time they tend to     harden and grow to the point of generating constant inflammation and     consequent pain. There is really no treatment, except for a     temporary anti-pain injection and the use of so-called “shock     waves”, which however have limited efficacy, some side effects and     complications, and a relatively high cost. In fact, in most cases     the only solution that is offered is the surgical removal of the     stones (calcium deposit) and this also has its problems because,     apart from the invasive effect of the intervention, the recovery and     rehabilitation periods are long and expensive. And there is no     guarantee that calcium deposits will not form again. We have found,     after numerous experiments, that the high level transporation of a     product based on AFA alga and AFA-phycocyanins extract, plus     Spirulina's phycocyanins extract, and some more specific ingredients     that we shall discuss, can be extremely effective in this field. The     first case we treated was done with the usual cyanobacterial     ingredients plus proteolytic and lipolytic enzymes, and there was     not only a radical pain and inflammation reduction, but also an     unexpected positive result on calcification. It was the case of a     lady who suffered from chronic Achilles's tendon degeneration, with     advanced calcification and inability to walk properly. His     specialist doctor had already set the appointment for a surgical     calcification removal. The patient underwent four sessions with the     transporation of the product described above. After four sessions     the pain had completely gone away, she returned to walk regularly,     even with high heels, and thus canceled the operation. At the same     time, the lady also made a magnetic resonance that showed how     calcification had been reduced by more than 60%. We have therefore     deduced that the AFA algae and phycocyanins extracts, in synergy     with specific enzymes, can promote the dissolution of calcium     deposits. This may in fact be mediated by anti-inflammatory action,     being known that calcium deposits are essentially modulated by     inflammation. The anti-inflammatory action of the molecules     contained in the algae has been shown to be useful also in reference     to this problem, as all the test patients reported significant     improvement in pain and mobility, even after the first treatment.

At this point, however, we decided to develop a more specific formula for such a painful and widespread problem, that has currently no satisfactory solution, adding to the product described above a mixture of specific minerals, endowed with the ability to generate the lysis and dissolution of calcium deposits. The list of the minerals added to the cyanobacterial formula includes, without being exhaustive, acetic acid, citric acid, EDTA (ethylenediaminetetracetic acid), sodium bicarbonate and sodium hydroxide.

This formula has been tested, the main ingredient still being the cyanobacterial algae and extracts, plus enzymes plus the above mentioned minerals. We have tested it in over 50 cases of more or less advanced calcification of the joints. Each of the participants in the study was asked to perform an ultrasound scan before the treatment, to evaluate the position and size of the calcification. Each participant with a proven calcification was then treated with the transporation of the above mentioned formula, and the result was verified through a further post-treatment ultrasound scan. In all 50 cases, on average after only four sessions and sometimes after 6 sessions, calcifications were completely dissolved, and the patients completely recovered the mobility of the joint. We believe that the transporation of the above-described formula can be an effective therapeutic solution of a previously unresolved problem that afflicts an ever-growing segment of the population. Here we report only two of the 50 cases that were recorded:

-   -   a) a 38-year-old man with a 15 mm (1.5 cm) long shoulder         calcification, that was completely removed after four sessions         of the trans-poration of the same formula, with the patient         recovering the full arm and shoulder functionality (FIG. 20);     -   b) a 65-year-old woman who had developed a calcification of her         shoulder many years before, which hindered her function and         constantly caused pain; she was treated with the trans-poration         of the formula described above, and the calcification was         completely dissolved (FIG. 21);

-   METABOLIC—We did some tests with interesting results on metabolic     dysfunctions such as lymphatic congestion (due to diabetes or aging)     and circulatory problems (varicose veins, heavy legs, etc.).     Phycocyanins (AFA-PCs or Spirulina PCs) also have anti-platelet     aggregation activity, and thus contribute to adequate blood flow,     thereby counteracting excessive blood density which is also     responsible, at least in part, for the most common circulatory     problems such as varicose veins. In addition, inflammation plays an     important role in such problems, and we have already indicated how     phycocyanins express their anti-inflammatory action much more     effectively if applied in situ through transporation. In addition,     circulatory problems are also related to the weakening of the     capillary arteries as well as, more generally, the laxity of the     parietal membranes of the blood vessels. Phycocyanins have a     significant stimulating activity of fibroblasts, which are also     involved in the production and maintenance of structures such as     collagen, elastin and more generally connective tissue, which are     essential for the health and tone of the blood vessels. Therefore,     AFA-PCs as well a Spirulina PCs, administered transdermally, can     play an essential role in the treatment of vascular problems.     Further contributions to the efficacy of the treatment were made     with the addition of proteolytic and lipolytic enzymes, which are     characterized by both a specific anti-platelet aggregation and     fibrinolytic activity. We treated two women, one younger (35) and     one older (65), with varicose veins in the legs. Both women, after     three treatments the first, after four treatments the second,     reported a visible reduction in the amplitude and depth of the     varicose veins, claiming to feel much better, with a feeling of leg     lightness.

-   SYSTEMIC—The possibility of achieving significant depths though high     level transporation of large molecules, thus potentially reaching     through local circulation the inner organs, opens up the possibility     of a systemic medical use of the methodology. For example,     phycocyanins have a significant anti-proliferative activity     (Kunte M. et al, 2017; Ravi M. et al., 2015); and our research group     has shown significant anti-proliferative activity for     AFA-phycocyanins from Klamath algae (Scoglio S. et al., 2016). It is     possible to hypothesize the possibility of affecting, through the     high intensity transporation of phycocyanins, organs affected by     malignant growth in order to inhibit such growth directly. The same     goes for other areas, such as the dissolution of gallstones and     kidney stones via the direct transporation on the specific areas.

As a first proof of this hypothesis/possibility, we treated a child with cystic fibrosis, who had already improved his condition through a nutritional supplement program with Klamath algae and different types of fungal enzymes. The parents agreed to submit the child to a series of sessions where the cyanobacterial product (Klamath algae, AFA-phycocyanincs extract, Spirulina's phycocyanins extract), enhanced by proteolytic and lipolytic enzymes, was applied via electroporation to the chest and the back of the child, to see if this could have an effect on the excessive lung congestion affecting the young cystic fibrosis patient. During each of the three sessions, the child expelled a large amount of lung catarrh, interpreted as a positive sign by both physicians and parents. This case preliminarily proves that the transporation of the above described product is able to penetrate so deep as to penetrate the organs, in the specific case the lungs and bronchial tubes.

REFERENCES

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1. The transdermal vehiculation of cyanobacterial algae, and/or extracts thereof, via electroporation, sonoporation, radio frequency, photoporation, or any other system using some type of force to allow for the opening of the water and ionic channels of the skin, for beauty and health enhancing purposes.
 2. The use, according to claim 1, of specific devices, including without being limited to, electrical currents or impulses, such as ionophoresis, iontophoresis, electrical muscle stimulation, and all forms of electroporation; vibratory currents, such as cavitation, ultrasound and all forms of sonoporation; laser, LED light and all forms of phototherapy; all forms of radio frequencies; thermal devices using heat; any other device capable of opening the skin's pores to allow for the penetration, at different levels of depth, of the large molecules.
 3. A preparation, according to claim 1, which contains the whole Klamath AFA algae and/or extracts thereof, and/or whole Spirulina or Spirulina's extracts, and/or any other cyanobacterial algae and/or cyanobacterial extracts, combined with any other substances.
 4. A preparation according to claim 2, whereby the other substances include, without being limited to, purified phycocyanins, purified phenylethylamine, proteolytic and lipolytic enzymes, hyaluronic acid, glycosaminoglycans, chlorophyll, carotenes, xantophylls (astaxantin, canthaxanthin, zeaxantin, lutein, licopene), polyphenols, polysaccharides, caffeine, vitamins, minerals and herbs.
 5. A preparation, according to claim 1, which is formulated in pharmaceutically acceptable vehicles and excipients, wherein all components of the preparation are suitable for administration by transporation methods.
 6. The use of a preparation, according to claim 1, applied transdermally via transporation to a subject for preventing, controlling, mitigating or treating aesthetic problems, such as wrinkles, cellulite, stretch marks, age spots, skin and muscle tone, breast tone and volume.
 7. The use of a preparation according to claim 1, applied transdermally via transporation to a subject for preventing, controlling or treating dermatological diseases, including, without being limited to, acne, eczemas, keloids, psoriasis, pimples, scars, wounds.
 8. The use of a preparation according to claim 1, applied transdermally via transporation to a subject for preventing, controlling, mitigating or treating muscle and osteo-articular diseases such as arthritis, joint inflammation, joint calcification, tendon inflammation and calcification, cartilage degenerations such as meniscus, osteoporosis.
 9. The use according to claim 8, whereby the preparation used for joint and tendons calcification also comprises specific calcium dissolving minerals, including, without being limited to, acetic acid, citric acid, EDTA (ethylenediaminetetracetic acid), sodium bicarbonate and sodium hydroxide.
 10. The use of a preparation according to claim 1, applied transdermally via transporation to a subject for preventing, controlling, mitigating or treating metabolic diseases such as lymphatic (diabetes, ageing) and circulatory problems (varicose veins, heavy legs, etc.), obesity, fat accumulation and metabolism.
 11. The use of a preparation according to claim 1, applied transdermally via transporation to a subject for preventing, controlling, mitigating or treating organ-centered degenerations, including, without being limited to, hepatitis, gallbladder and kidney stones, intestinal and respiratory problems.
 12. The use of a preparation according to claim 1, wherein the quantity of preparation applied transdermally is comprised from 0.1 to 500 gr and preferably of approximately 1 to 10 grs., according to the size of the area to be treated.
 13. The use of a preparation according to claim 1, wherein the composition is in the form of a powder to be dissolved in distilled water, or in the form of a creme, gel or liquid suitable for administration by transporation methods.
 14. The use according to claim 1, wherein the subject is an animal or human subject. 